POTENT AND SELECTIVE-INHIBITION OF HUMAN NITRIC-OXIDE SYNTHASES - INHIBITION BY NON-AMINO ACID ISOTHIOUREAS

S-Ethylisothiourea was a potent competitive inhibitor of human nitric oxide synthase (NOS), with K-i, values of 17, 36, and 29 nM for the in ducible (i), endothelial (e), and neuronal (n) isozymes, respectively. Unlike some potent inhibitors of NOS, no time dependence was observed . S-Ethylisothiourea was not a detectable substrate for eNOS. S-Ethyli sothiourea was also a potent inhibitor of mouse iNOS (K-i value of 5.2 nM), and its binding perturbed the spectrum of iNOS consistent with i ts altering the environment of the bound heme. The optimum binding of S-ethyl- and S-isopropylisothiourea relative to 70 other analogs sugge sted that these alkyl substitutions fit into a small hydrophobic pocke t. Most isothioureas were 2-6-fold selective for the human iNOS (K-i f or iNOS versus K-i for eNOS), with one being 19-fold selective. The cy clized mimics of S-ethylisothiourea, 2-NH2-thiazoline, and 2-NH2-thiaz ole, were also competitive inhibitors of human NOS. A third structural class of inhibitors, bisisothioureas, were, in general, the most sele ctive in their inhibition of human iNOS. '-(1,3-Phenylenebis(1,2-ethan ediyl))bisisothiourea was 190-fold selective (K-i value of 0.047 mu M against iNOS versus 9.0 mu M against eNOS). These results demonstrate that potent and selective inhibition of human NOS isozymes is achievab le.